枯草芽孢杆菌α-淀粉酶基因的克隆及表达

Cloning and expression of α amylase gene from Bacillus subtilis HN503

采用鸟枪法克隆到了枯草芽孢杆菌α 淀粉酶基因 .试验证实 ,已报道的α 淀粉酶的基因大小为 2kb左右 ,因此回收 3～ 7kb的目的片段较适宜 .对质粒去磷可防止自连 ,并能显著提高外源片段的连接效率 ,采用扩大酶连的方法有助于酶连产物的进一步提高 .高效感受态比普通感受态构建基因文库效率要高 2个数量级 .高效、快捷的鉴定α 淀粉酶的筛选培养基是克隆α

The α amylase gene of Bacillus subtilis HN 503 was successfully cloned by shotgun method. The experiments showed that 3～7kb target DNAs were extracted according to the report that the α amylase gene is 2kb or so. The rate of foreign DNA ligation was improved with plasmid KS dephosphorylated by CIP and the rate was increased again by enlarged ligation. Gene library was constructed by high efficiency competence cell, which was 100 times effectiver than by that of CaCl 2 method. High efficiency, fast and easy way to identity the α amylase was necessary.